The role of sperm inner acrosomal membrane proteins in acrosomal biogenesis and fertilization

受精是发育的基石，精子和卵子这两个分化的细胞为胚胎发生和以后的发育奠定了基础，配子的能力是必不可少的。然而，关于精子-卵子相互作用导致合子发育的分子和机制，只有基本的知识可用。考虑到30%-50%的男性因素不育症病例没有已知原因，这一点得到了强调。我们了解这些机制的策略是分离和分子解剖参与受精过程的精子隔室(例如顶体和核周膜)。这些蛋白质的同一性为我们提供了了解这一间隔的重要性的线索，并使我们能够设计特定的探针(例如抗体、野生型和突变型重组体和竞争性抑制物)来测试这些蛋白质在受精和/或精子发生过程中的预测功能和机制。利用这种方法，在NSERC发现基金的授权下，我和我的学生实习生研究了顶体内膜(IAM)的功能。这种精子成分在受精过程中起着关键作用，因为它被顶体反应暴露出来，然后将精子牢固地结合到卵子的透明带(ZP)上，这是随后IAM指导的精子穿透细胞外基质的必经步骤。在我们的研究之前，IAM的蛋白质还没有被探索过。我们已经鉴定并鉴定了以下IAM蛋白：IAM38，与ZP结合的受体；MMP2，潜在的ZP穿透酶；PRACESIN，潜在的MMP激活酶；SPACA1，可能组织这些蛋白并将其附着到IAM的跨膜蛋白；SubH2Bv，Kapa和Kap？，它们形成了一个复合体，可能在精子发生过程中将顶体小泡引导到细胞核。本提案旨在验证这些蛋白质的潜在功能，并阐明它们的作用机制。所获得的知识无疑将有助于更好地理解精子不育缺陷的起源及其诊断。它还可以帮助改进辅助生殖技术的应用，并设计一种避孕药具。

Fertilization is the cornerstone of development where the two differentiated cells, sperm and egg set the stage for embryogenesis and beyond, for which competency of the gametes is essential. Yet only rudimentary knowledge is available on the molecules and mechanisms involved in sperm-egg interactions leading to zygotic development. This is underscored in consideration that 30-50 % of male-factor infertility cases have no known cause. Our strategy in gaining an understanding of these mechanisms is to isolate and molecularly dissect sperm compartments (e.g., acrosome and perinuclear theca) that are involved in the fertilization process. The identities of the proteins provides clues to the compartment's significance and allows us to design specific probes (e.g., antibodies, and wild type and mutant recombinants and competitive inhibitors) to test the predicted functions and mechanisms of these proteins during fertilization and/or spermiogenesis. Utilizing this approach, under the mandate of the NSERC Discovery Grant, my student trainees and I have investigated the function of the inner acrosomal membrane (IAM). This sperm component plays a critical role during fertilization as it is exposed by the acrosome-reaction and then binds the sperm firmly to the zona pellucida (ZP) of the egg, a compulsory step for subsequent IAM-directed sperm penetration of this extracellular matrix. Before our study the proteins of the IAM were unexplored. We have identified and characterized the following IAM proteins: IAM38, binding receptor to ZP; MMP2, potential ZP penetrating enzyme; proacrosin, potential MMP activating enzyme; SPACA1, trans-membrane protein which potentially organizes and attaches these proteins to the IAM; SubH2Bv, KAPa and KAPß which form a complex and potentially direct the acrosomic vesicle to the nucleus during spermiogenesis. The present proposal intends to validate the potential functions of these proteins and elucidate their mechanisms of action. The knowledge gained would unquestionably lead to a better understanding of the origins of sperm infertility defects and their diagnosis. It could also serve to improve application of Assisted Reproductive Technologies and to design a contraceptive.